This research proposal is submitted in response to the NIH announcement (NOT-OD-09-058) on availability of Recovery Act Funds for Competitive Revision Applications. The currently funded parent grant (UOI AA014095) to this competitive revision application is a research component of the NIAAA supported INIA- Stress Consortium, which is focused on stress-ethanol interactions in relation to excessive drinking behavior. Specifically, the project aims to examine mechanisms by which stress associated with repeated cycles of chronic ethanol exposure and withdrawal promotes excessive ethanol drinking and increased vulnerability to relapse associated with ethanol dependence. Utilizing a mouse model of ethanol dependence and relapse/excessive drinking, major emphasis is focused on studying the role of CRF. This competitive revision proposal is designed to extend and enhance our efforts to elucidate adaptive changes in CRF activity within brain reward and stress pathways that are associated with ethanol dependence and escalation of drinking behavior. While studies in the parent grant focus on neuroadaptive changes in CRF in the mouse model of ethanol dependence and relapse drinking, these studies are limited to measuring brain regional changes in mRNA (by quantitative real-time PCR (qRT-PCR) analysis) and peptide content (by ELISA) of CRF itself. Studies proposed in this competitive revision application will extend and complement work conducted in connection with the parent grant by focusing on transcriptional and protein changes in CRFI and CRF2 receptors using the same ethanol dependence and relapse drinking model. Further, in addition to analysis of brain CRF1 and CRF2 receptor mRNA and protein by qRT-PCR and immunoblotting, in situ hybridization procedures will be used to gain greater anatomical resolution of neuroadaptive changes in CRF function associated with the model of ethanol dependence and excessive drinking. Overall, the goal is to provide a more comprehensive analysis of adaptive changes in functioning of the CRF stress neuropeptide system within brain reward and stress pathways that result from stress associated with ethanol dependence and contribute to excessive drinking behavior and the development of alcoholism. PUBLIC HEALTH RELEVANCE: While stress has been viewed as an important contributing factor to alcohol abuse and alcoholism, the interaction between stress and ethanol drinking behavior is not well understood. This is especially true when one considers the role/impact of stress on ethanol self-administration in the context of dependence. The overall goal of this research project is to gain a better understanding about the role of stress in promoting excessive ethanol drinking and increased vulnerability to relapse associated with ethanol dependence.